Vitamin D deficiency is associated with the development of some cancers and in vitro 1,25-dihydroxyvitamin D (1,25D) reduces cell proliferation. We suggest that modification of tissue specific immune responses, as a consequence of local synthesis of 1,25D, may be key. To assess the impact of serum 25D on the risk of bladder cancer we conducted a systematic review. To test our hypothesis, expression of vitamin D signalling components and the synthesis of 1,25D were examined in human bladder epithelial cell lines (T24/83 and RT4). A search of Embase, Web of Science, Medline and Cochrane library (AprilMay 2016) identified 287 citations. Following title and abstract review by two reviewers seven full papers were appraised. Studies varied in the number of participants (1121125) and point of vitamin D measurement (pre-diagnosis, diagnosis, or follow-up). Low vitamin D levels were associated with bladder cancer risk in five of the seven studies. Higher vitamin D levels also correlated with better survival and outcomes. The vitamin D receptor and 25-hydroxyvitamin D 1α-hydroxylase (CYP27B1; 1α-OHase) mRNA and protein were expressed by both cell lines. 24-hydroxylase (24-OHase; metabolises 1,25D) mRNA was almost undetectable in unstimulated cells but was increased significantly by 1,25D (10 nM, 324 h; P<0.05). 24-OHase was also induced by 25D (100 nM, 624 h; P<0.05) indicating 1α-OHase activity. Synthesis of 1,25D was confirmed by EIA. Cathelicidin mRNA was induced by 1,25D and 25D in RT4 cells (10 nM/100 nM, 6 h; P<0.05). These data demonstrate that bladder cancer risk correlates with low serum 25D levels. Transitional epithelial cells express functional vitamin D signalling and are able to synthesize sufficient 1,25D to stimulate a local immune response. We propose that in order to initiate a cell-mediated immune response to malignancy adequate levels of serum 25D are required for synthesis of 1,25D by bladder epithelial cells.